Prokaryotic and eukaryotic cells contain multiple types of RNA at least some of which can be characterized by different chemical constituents at their 5′ ends. Within all cells, RNA polymerase initiates synthesis of RNA with a 5′ terminal nucleotide having a 5′ triphosphate. 5′ monophosphate nucleotides are then successively added to the 3′ end. However, for eukaryotes, RNA destined to become messenger RNA (mRNA) is rapidly capped with a m7G nucleotide linked to the 5′ terminus via a 5′ triphosphate linkage. This modification is a result of the capping enzyme. In contrast, the mRNA of bacteria and archaea maintain their 5′ triphosphate. In all kingdoms of life, ribosomal RNA (rRNA) makes up the vast majority of the cell's RNA but its 5′ end is generated by endonucleolytic cleavage to leave a 5′ monophosphate terminus. Furthermore when RNA is degraded in the cell by RNases, the 5′ ends are either 5′ monophosphate or 5′ hydroxyl groups.
When analyzing the sequence and quantity of specific RNAs, it is desirable to remove rRNA and degraded RNA from the complex mixture of RNAs. There are currently a number of procedures for rRNA depletion, but they all suffer some shortcomings. For example, depletion methods commonly require the hybridization of DNA oligonucleotides complementary to rRNA and removal of the hybrid molecules. This requires customizing the DNA sequence to match the rRNA. It also requires a priori knowledge of the ribosomal sequence. Another procedure that has been used has been the specific enzymatic degradation of 5′ monophosphate RNA that would include rRNA, however this enzymatic reaction has proved inefficient and leaves substantial rRNA and degraded RNA in the mixture.